The mechanisms linking nutrient intake to altered bone formation are poorly understood. Although most research has focused on the calcium-PTH-Vitamin D axis, our laboratory has a longstanding interest defining the mechanisms utilized by enteric hormones to modulate bone turnover. During the conduct of our research, we observed that nutrients not only affect bone turnover by indirect mechanisms (e.g. by modulating release of bone active hormones such as IGF-1, GLP-2, GIP or by modulating the activity of the innervation in bone) but also exert direct effects on bone marrow stem cells (BMSCs). Although it has been known for some time that nutrients can modulate osteoblastic and osteoclastic activity, direct nutrient effects on BMSCs have not been demonstrated. Preliminary data presented in this proposal support a role for direct, receptor-mediated nutrient effects on BMSC proliferation, specifically: (1) BMSC do not grow or differentiate in growth mediums with low amino acid concentrations; (2) BMSC express extracellular L-type amino acid sensors [the calcium sensing receptor (CasR) and the taste receptor (T1 R1/T1R3)]; (3) aromatic amino acids increase intracellular calcium; (4) aromatic amino acids and glutamate activate the MAPK signaling pathway; (5) expression and function of specific extracellular amino acid sensors vary in an age-dependent manner; and (6) aging C57BI6 mice fed a low protein diet lose bone and this loss is prevented by dietary supplementation of aromatic amino acids. We hypothesize that direct nutrient effects on BMSCs play an important role in preserving bone mass. This proposal focuses on defining the contribution of direct nutrient effects on BMSCs to bone formation using interrelated in vivo and in vitro studies. Therefore, the specific aims of the proposal are: (1) To define the molecular mechanisms involved in direct amino acid induced BMSC/Osteoprogenitor cell activation in vitro; and (2) To define the effect of dietary nutrients (protein) on bone mass in vivo. Findings from the proposed experiments should help reveal and define the mechanistic links between nutrient ingestion and bone formation and result in new targets for therapeutics, particularly those targeted to age-dependent alterations in bone formation. This project also will explore the interactions between amino acid effects on BMSC with the fat and muscle derived hormones, leptin and IGF-1 and the cytokine SDF-1 and thus provides a unique opportunity to examine nutrient effects on bone in a comprehensive manner.